This application is submitted in response to Grand Opportunities (RC2): Engineered Nanomaterial Environmental Health and Safety. The increasing use of engineered nanomaterials in industrial and medical applications is expected to increase both unintended environmental or occupational exposures and intended medical or direct consumer exposures, but the impact of such exposures on human health is unclear. The potential toxicity or biocompatibility of engineered nanomaterials is governed by the cellular interactions and fate of the particles, which dictate the cellular response and ultimately determine the impact on human health. The cellular interactions and subsequent response of the cells are governed by the physical and chemical properties of the particles, but the relationships between particle properties and these cellular processes are far from being understood. Furthermore, the properties of nanomaterials are likely to be modified by the environment, such as ambient air, but these changes are also unclear. The purpose of this proposal is to identify relationships between distinct properties of airborne engineered nanomaterials and their cellular interactions, fate, and response in alveolar epithelial cells at the air- liquid interface with the goal of supporting predictive evaluation of inhaled nonmaterial's toxicity or biocompatibility. Airborne nanomaterials that enter the respiratory tract are likely to be deposited in the alveolar region, where alveolar epithelial cells are found. These cells provide a vulnerable target for particles that escape the first line of defense by the alveolar macrophages. Accumulating observations indicate that nanomaterials are likely to be presented to alveolar cells in vivo as individual particles or small nanoscale aggregates, which differ from the larger particles in their ability to interact with the cells. We will establish methods for realistic exposures to well-defined monodispersed nanomaterials in ambient air for delineating relationships between distinct properties that are relevant to airborne particles and their impact on alveolar epithelial cells at the air- liquid interface. Size exclusion methods will ensure exposures to individual nanoparticles or small nanoscale aggregates, as they are likely to be presented to the cells in vivo. Building on our experience in quantitative fluorescence imaging with single molecule sensitivity, combined with molecular biology techniques, we will investigate the cellular interactions and fate of one nanoparticle or nanoscale aggregate at a time, delineating cellular processes that are relevant to the properties of the individual nanoparticle and the exposures in vivo. We propose to focus on surface modified and unmodified titania and amorphous silica nanoparticles, which have been widely used in diverse applications and pose a significant source for potential airborne exposures. Using analytical and physical chemistry methods, the properties of the particles, collected at the air-liquid interface, will be characterized. This information will derive changes that occur to nanomaterials in ambient air and delineate properties that are relevant to airborne nanoparticles and their cellular interactions and impact in vivo. Together, our studies will gain critical new relationships between properties of airborne nanomaterials and their cellular interactions, fate and response, supporting predictive evaluation of toxicity or biocompatibility of inhaled nanomaterials. The new information will have a large scale impact by guiding preventative approaches that will protect human health from adverse effects of engineered nanomaterials and the design of safe nanomaterials for new industrial and medical applications. PUBLIC HEALTH RELEVANCE: The increasing use of engineered nanomaterials in industrial and medical applications is expected to increase both unintended environmental or occupational exposures and intended medical or direct consumer exposures, but the impact of such exposures on human health is unclear. The potential toxicity or biocompatibility of engineered nanomaterials is governed by the cellular interactions and fate of the particles, which dictate the cellular response and ultimately determine the impact on human health. These cellular interactions and subsequent response of the cells are governed by the physical and chemical properties of the particles, but the relationships between particle properties and these cellular processes are far from being understood. Our research will address the specific call and public health needs by identifying critical new relationships between properties of airborne nanomaterials and their cellular interactions, fate and response, supporting predictive evaluation of toxicity or biocompatibility of inhaled nanomaterials. The new information will have a large scale impact by guiding preventative approaches that will protect human health from adverse effects of engineered nanomaterials and the design of safe nanomaterials for new industrial and medical applications.